# TAMPA BAY Numeric Nutrient Criteria: Task 1 - TN and TP Concentrations

Letter Memorandum

Prepared for:

Tampa Bay Estuary Program

Prepared by:

Janicki Environmental, Inc.

16 February 201116 February 2011 1 

### FOREWORD

This letter memo was produced in partial fulfillment of Purch ase Order #6584, TBEP Contract T-07- 01 - Development of Numeric Nutrient Criteria for Tampa Bay, Ta sk 1.16 February 2011 2 

### ACKNOWLEDGEMENTS

We wish to thank the partners of the Tampa Bay Estuary Program a nd members of the Tampa Bay Nitrogen Management Consortium for the numerous conversations pro viding direction and insight into concerns regarding numeric nutrient criteria establishmen t and appropriate methodology for developing the proposed criteria.16 February 2011 3 

### EXECUTIVE SUMMARY

The Tampa Bay Estuary Program (TBEP) and the Tampa Bay Nitrogen M anagement Consortium (TBNMC) have recommended to EPA numeric nutrient criteria for Tampa Bay as segment-specific annual total nitrogen and total phosphorus loads. EPA ha s noted its intention to express the numeric nutrient criteria for Tampa Bay as TN and TP concen trations. This document provides segment-specific TN and TP concentrations consistent with the TN and TP loads recommended as numeric nutrient criteria.
Establishment of numeric nutrient criteria is dependent on an u nderstanding of the limiting nutrient within the water body of concern. For Tampa Bay, extensive data exist for evaluation of which nutrient, nitrogen or phosphorus, is limiting. Ambient wat er quality data strongly indicate that Tampa Bay is nitrogen limited, and this is supported by the resul ts of nutrient addition bioassays.
Previous efforts by the TBEP have developed strong relationships betw een nutrient supply to Tampa Bay and resultant chlorophyll a concentrations in the bay, and between chlorophyll a concentrations and light availability for seagrasses. Thus, management actions have focused on controlling nitrogen loads to Tampa Bay, with measureable succ ess as expressed by increases in seagrass acreage. The relationships are between nitrogen loads an d chlorophyll a , however, not nitrogen concentrations in the bay and chlorophyll a

. Previous efforts to link in-bay nitrogen concentrations to chlorophyll a concentrations have not produced sufficient explanatory relationships for use in nit rogen concentration target setting. The current effort, driven by EPA’s intention to express numeric nut rient criteria as in-bay concentrations, examines several methods to link nutrient concentr ations to chlorophyll a concentrations and/or nutrient loads.
The final method selected for establishing proposed concentratio n numeric nutrient criteria for nitrogen and phosphorus was the reference period approach, b ased on the following rationale. Segment-specific chlorophyll a targets (values at this level or below indicate desirable conditi ons) have been established previously (Janicki and Wade, 1996). T hese targets were based on a 1992- 1994 reference period. In 2001, a protocol for assessing wh ether the Tampa Bay segments were achieving these targets was developed (Janicki Environmental, 2001). This protocol, referred to as the Decision Matrix approach, considered the year-to-year vari ability in chlorophyll a concentrations and arrived at segment-specific chlorophyll a thresholds (values above this level indicate undesirable conditions). The threshold was the sum of the chlorophyll a target and 2X the standard error of the long-term chlorophyll a concentrations.
Using the same approach, the segment-specific annual geometric mean nit rogen and phosphorus concentrations of the 1992-1994 period were derived as co mmensurate with the chlorophyll a targets. These concentrations were increased by one standard devi ation (as derived from 1992- 2009 data) to develop the proposed concentration numeric nu trient criteria, which serve as the threshold concentrations in the bay.16 February 2011 4

The following conclusions are drawn from the analyses completed for this effort:

• Tampa Bay is nitrogen-limited as indicated by both ambient TN:T P ratios and nutrient addition bioassays.

• There is no discernable relationship between TN loadings and i n-bay TN concentrations or between TP loadings and in-bay TP concentrations in any bay se gment. This is not because loadings do not affect in-bay concentrations, but because va rious other confounding factors, for which sufficient data are not available, play a role i n relationships between loadings and concentrations.

• On a monthly time scale, the relationships between either TN concen trations or TP concentrations and chlorophyll a concentrations do not explain a significant proportion o f the variability in the chlorophyll a concentrations to support development of concentrationbased numeric nutrient criteria in any bay segment.

• There are differences in TN concentrations and TP concentrat ions, particularly in Hillsborough Bay and Middle Tampa Bay, in those years when the chl orophyll a concentration thresholds are met when compared to those observe d in years when the chlorophyll a concentration thresholds are not met. However, there is a great deal of variability within the data obtained in either group of yea rs and these differences are not recommended as the basis for the establishment of concentration- based numeric nutrient criteria for Tampa Bay.

• The reference period approach is recommended for the establishmen t of concentrationbased TN and TP criteria for Tampa Bay. The segment-specific a nnual geometric mean nitrogen and phosphorus concentrations of the 1992-1994 pe riod were increased by one standard deviation (as derived from 1992-2009 data) to develo p the proposed concentration numeric nutrient criteria. These criteria are:
- Old Tampa Bay TN=0.93 mg/L TP=0.31 mg/L - Hillsborough Bay TN=1.01 mg/L TP=0.45 mg/L - Middle Tampa Bay TN=0.87 mg/L TP=0.29 mg/L - Lower Tampa Bay TN=0.74 mg/L TP=0.10 mg/L.

The criteria referenced above should be assessed as an annual geo metric mean from long-term, monthly water quality monitoring stations currently used in th e state’s chlorophyll a threshold assessments under the Tampa Bay Reasonable Assurance determination. T he assessment of TN and TP concentrations attainment should only occur when chlo rophyll a thresholds are exceeded within a bay segment, and should coordinate with current regu latory assessments under the FDEP RA determination and EPA TMDL for TN loads in Tampa Bay. Fur ther, compliance assessments should be conducted over five-year time frames, with no more tha n two consecutive years being greater than these established criteria if chlorophyll a thresholds are also exceeded during the same time period. This approach is analogous to the chlorophyll a threshold assessments currently being conducted under the regulatory requirements for the FDEP RA det ermination and EPA TMDL for Tampa Bay.16 February 2011 5



### 1.0 Introduction and Objective

The Tampa Bay Estuary Program (TBEP) and the Tampa Bay Nitrogen Ma nagement Consortium (TBNMC) have recommended numeric nutrient criteria to U.S. En vironmental Protection Agency (EPA) for Tampa Bay (TBNMC, 2010). The criteria, as p roposed to EPA, are segment-specific (Figure 1) and are expressed as annual total nitrogen (TN) an d total phosphorus (TP) loads. These TN and TP loads are those for the reference period of 1992- 1994, as discussed in the March 8, 2010 comments to EPA. However, EPA has informed the TBEP that it intends to express the numeric nutrient criteria for Tampa Bay as TN and TP concen trations.
The objective of this task is to develop segment-specific TN and T P concentrations consistent with the TN and TP loads recommended as numeric nutrient criteria b y the TBEP and TBNMC. This is in keeping with recognition of the importance of maintaining c onsistency with existing management goals and specifically with the recent load allocat ions to comply with the existing TMDL for Tampa Bay.

The following provides a discussion of nutrient limitation, description of the analyses completed and the results of each analysis, and the recommended TN and TP numeric nutrient criteria, expressed as concentrations, for each of the four mainstem bay segments.



### 2.0 Nutrient Limitation

The establishment of numeric nutrient criteria depends upon kn owledge of the nutrient most likely limiting in the waterbodies of concern. Three major factor s control whether nitrogen or phosphorus is more likely to be limiting (NRC, 2000):

• the N:P ratio in external nutrient inputs;

• the preferential loss from the photic zone of nitrogen or p hosphorus due to biogeochemical processes such as denitrification, sedimentation, or absorption of phosphorus; and

• the amount of nitrogen fixation.
Marine systems, including estuaries, are generally considered nitr ogen limited (Thomas, 1970a,b; Ryther and Dunstan, 1971; Boynton et al., 1982; Smith, 1984; Howarth, 1988, 2008; Howarth et al., 1988a,b; Nixon et al., 1996; Howarth and Marino, 2006; Chapra, 1997; National Research Council, 2000;), although there may be times and locations wh en phosphorus limitation may occur (Conley, 2000; Conley et al., 2009; Malone et al., 1996).16 February 2011 6 Figure 1. Tampa Bay and its four major bay segment s. 

## Old Tampa **Bay**

**Hillsborough Bay** 

**Middle Tampa Bay** 

**Lower Tampa**  **Bay**

Gulf of Mexico16 February 2011 7

Since nitrogen is considered the most likely limiting nutrient in est uarine systems, it has been identified as the primary nutrient of concern in estuarine eco systems nationwide (Smith, 1984; NRC, 1993). As noted in Correll (1999), however, since estua ries are part of the transition zone between the open ocean and the phosphorus supplied from the land , it is possible that both phosphorus and nitrogen may be limiting in estuaries, dependent upon the time of year, location in the estuary, and nutrient supplies. Ryther and Dunstan (1971 ) noted the change from phosphorus limitation in freshwaters to nitrogen limitation in near-sh ore marine waters, although Hecky and Kilham (1988) indicated that the extent and severity of marine n itrogen limitation has not been conclusively determined, with other studies reporting estuarine phosp horus limitation in the spring and nitrogen limitation in the summer and fall (Fisher et al. , 1992; Lee et al., 1996). Depending upon the relative rates of nitrogen and phosphorus supply, the limitation has been found to shift between nitrogen and phosphorus in coastal lagoons in the nort heastern US (Taylor et al., 1995). It has also been documented that residence times play a significant role in determining the estuarine responses to nutrient loads ((Monson et al., 2002; Hagy et al ., 2000; Borsuck et al., 2004; Boynton and Kemp, 2008).

2.1 Methods to Determine Limitation
There are two general methods that have been used to define whic h nutrient is limiting in a water body. They include:

• a method that depends upon ambient water quality data collect ed over a wide range of environmental conditions, and

• a method that involves experimental manipulation of nutrient co nditions, either in the laboratory or in situ

. 2.1.1 Ambient Water Quality Data Methods
This method depends upon a metric typically used to evaluate nutr ient limitation: the nitrogen to phosphorus ratio (N:P ratio). The N:P ratio indicati ve of balanced conditions is typically taken to be 16:1 (molar), based on the work of Redfield (1934, 1958). This N:P ratio was based on the elemental composition of algae, under both laboratory and na tural conditions. When N:P ratios are greater than 16:1 in a system, this is indicative of phosphorus limitation. However, there may be considerable variation in this ratio within an algal cul ture, dependent upon cell division status, light conditions, and precedent conditions (Correll, 1999; Corr ell and Tolbert, 1962; Terry et al., 1985).
Molar ratios of N:P are easily determined from water quality mo nitoring data. In freshwater systems, the N:P ratio is usually higher than 16:1, indicati ng that phosphorus is usually most limiting to primary production in these ecosystems (Schindler, 197 7; Elser et al., 2007). This becomes evident by examining the concentrations of the forms of ni trogen and phosphorus that are available for algal uptake. Little if any dissolved inorganic phosphorus (DIP) is generally found in relatively productive freshwaters while measurable concentrations of dissolved inorganic nitrogen (DIN) remain.
Most marine systems are nitrogen limited because there are relatively low concentrations of dissolved inorganic nitrogen compared to dissolved phosphorus. Since Redfield’s observations were published, research has shown that ratios from 10:1 to 20:1 for N:P are typically found in16 February 2011 8

estuaries (Parsons et al., 1984). Howarth (1988) observed th at the correlation between nitrogen and the primary production was better for estuaries that rec eived nutrient concentrations with smaller N:P ratios than the one studied by Redfield. Several studi es have led to the conclusion that estuaries receiving nutrient concentrations with high N:P ra tios were limited by phosphorus and only those with low ratios are limited by nitrogen (Boynton et al., 1982). Boynton et al. (1982) and Howarth (1988) compiled data on the ratio of inorganic nitrogen to phosphorus in a variety of estuaries. Of the 27 studied by Howarth, 22 had N:P ratios below the Redfield ratio and may have been nitrogen limited. Because phytoplankton can assimilate some organ ic nutrient forms and all forms are relatively labile, it is useful to examine the ratio of t otal nutrient concentrations (TN:TP).
Reductions of nutrient levels in a water body will usually resu lt in reduction in algal growth. Reducing phosphorus, however, will have no effect unless the reduc tion results in an N:P ratio greater than 16:1. Phosphorus would then become the limiti ng nutrient. In contrast, a reduction of nitrogen concentrations will result in a reduction of prima ry productivity when the ratio is less than 16:1. There are exceptions to this general rule. Some coastal ar eas are phosphorus limited due to strict phosphorus control measures or natural conditions an d some freshwaters are nitrogen limited due to natural sources of phosphorus.

2.1.2 Experimental Methods
Experimental manipulation of nutrient conditions, either in the lab or in situ, typically involves nitrogen and phosphorus additions to either a test alga or a phytoplankton assemblage singularly and in combination. The responses to the additions determin e the limiting nutrient. If growth is found only during nitrogen addition, nitrogen-limitation is indicated. Conversely, if growth is found only during phosphorus addition, phosphorus-limitat ion is indicated.

In situ methods have included:

• limnocorrals or bags in which nutrient additions are made a nd resultant growth responses are measured (Shapiro, 1980; Lynch and Shapiro, 1981; Havens a nd DeCosta, 1986; Perez et al., 1994);

• mesocosm studies in which water is collected and placed in separa te containers or enclosures for application of separate treatments over multipl e day time scales (Oviatt et al., 1986; Taylor et al., 1995); and

• whole-lake studies performed on entire lakes or portions of lakes separated by curtains (Schindler, 1974, 1975).
In Florida, as part of its TMDL process, the Florida Departmen t of Environmental Protection (FDEP) attempts to identify the limiting nutrient(s) in impaired waterb odies. The TMDL for a specific waterbody specifies the maximum amount of the limiting nutrient tha t may enter the waterbody, with this limitation being defined with the aim of improving w ater quality. If the N:P ratio does not clearly suggest the limiting nutrient, TMDLs for both nitrogen and phosphorus are typically defined. The primary method for determining the limiting nutrient employed by the FDEP is use of existing water quality data to derive ambient N:P ratios, but more comp licated methods, including field tests and laboratory algal growth potential bioassays, have been employed. Per FDEP guidelines, receiving waters with ratios less than 10:1 (molar) are consider ed nitrogen limited, ratios of greater than 30:1 (molar) indicate phosphorus limitation, and ra tios of 10-30:1 (molar) indicate colimitation (FDEP, 2002).16 February 2011 9

2.2 Confounding Factors
Determination of the limiting nutrient based solely on N:P rat ios estimated from water quality data or from experimental uptake rates should be performed with consi deration of potentially confounding effects. Algal cell interior N:P ratios and uptake rates may vary due to:

• cell division status (Correll and Tolbert, 1962),

• light intensity or light quality (Wynne and Rhee, 1986),

• light and temperature (Jahnke et ., 1986), and

• P deprivation and then subsequent availability (Sicko-Goad an d Jensen, 1976).

Nutrient limitation in freshwaters, which are typically consi dered to be phosphorus limited, can vary seasonally. Summer nitrogen limitation in lakes can occur when photic zone inorganic nutrients are low (Elser et al., 1990). It has also been demon strated that some estuaries show seasonal shifts in limitation (D’Elia et al., 1986; McComb et al., 1981; Conley, 2000). The best available information should be used to determine the limiting n utrient of a system before management decisions are made with the objective of improved water qu ality via nutrient load control.

2.3 Nutrient Limitation in Tampa Bay
Nutrient limitation in Tampa Bay has been examined using both the N:P ratio method and nutrient addition bioassays.

2.3.1 TN:TP Ratios in Tampa Bay
The average TN:TP ratios for the segments of the TBEP, both by weight and molar, were determined based on ambient water quality data, and are presen ted in Table 1 for the period 1981 to 2009. The ratios were calculated by first calculating the monthly ratio for each segment based on data collected by the Environmental Protection Commission o f Hillsborough County (EPCHC). The mean value of these monthly values within a year was calculated and the mean of these annual values was calculated.

Table 1. Annual mean TN and TP concentrations and TN:TP in TB EP segments (1981-2009).

Bay Segment TN (mg/L) TP

(mg/L) TN:TP

(Weight) TN:TP

(Molar) Old Tampa Bay 0.71 0.27 3.5 7.7 Hillsborough Bay 0.79 0.45 2.1 4.6 Middle Tampa Bay 0.63 0.27 2.9 6.3 Lower Tampa Bay 0.48 0.13 4.8 10.5

All segments except Lower Tampa Bay have molar N:P ratios less than 1 0:1, while that for Lower Tampa Bay is just slightly greater than 10:1. According to th e FDEP guidelines (FDEP, 2002), all segments would therefore be considered nitrogen-limited. Whil e nitrogen limitation is indicated in all segments, the nutrient that is most limiting can vary seasonally ( Malone et al

. , 1996; Conley et16 February 2011 10

al., 2009), so that areas that are generally nitrogen limit ed may by phosphorus-limited at times. In addition to nutrient limitation, phytoplankton growth ma y also be light-limited during certain parts of the year (Pennock and Sharp, 1994).

Seasonal variation in nutrient limitation has been observed in other waterbodies (Fisher et al., 1992; Lee et al., 1996; Malone et al, 1996; Conley et al., 2 009). Season-specific TN:TP ratios were also estimated based on the 1981-2009 data (Table 2). These estimates continue to support the conclusion that Tampa Bay is nitrogen-limited.

Table 2. Seasonal mean TN:TP ratios in TBEP segments (1981- 2009 ).

Bay Segment Dry Season Wet Season
TN:TP (Weight) TN:TP

(Molar) TN:TP

(Weight) TN:TP

(Molar) Old Tampa Bay 3.6 8.0 3.2 7.1 Hillsborough Bay 2.2 4.9 1.9 4.2 Middle Tampa Bay 3.0 6.6 2.6 5.7 Lower Tampa Bay 5.1 11.3 4.2 9.2

2.3.2 Nutrient Addition Bioassays in Tampa Bay
The City of Tampa Bay Study Group routinely performed bioassays th roughout Tampa Bay during 1993-2009 as part of the evaluation of the effects of discha rge from the H.F. Curren wastewater facility (Johansson, 2009 – 

### Attachment 1 ). Bioassays in late winter and late summer have been conducted for most of this period. The results of these 152 b ioassays have supported that nitrogen is the primary limiting nutrient in the bay, with no results sho wing phosphorus as the limiting nutrient in the bay, including within that portion of t he bay which receives the wastewater effluent. Therefore, the discharge phosphorus limitation is not required for this facility.
It is concluded that the four mainstem bay segments in Tampa Bay a re nitrogen-limited. This conclusion contributed to the development of TN loading tar gets for Tampa Bay (Janicki and Wade, 1996), without consideration of TP loading targets in th e original target setting effort. Since EPA is considering developing both TN and TP numeric nutrient crit eria for Tampa Bay, the proposed numeric nutrient criteria expressed as TP loads were based on t he same time period used to develop the proposed TN loading numeric nutrient criteria (c omments submitted to EPA (TBNMC, 2010). Table 3 presents the recommended protective nutrient load s established by the TBNMC and proposed to EPA.16 February 2011 11

Table 3. Protective nutrient loads for the Tampa Bay estuary established by the Tampa Bay Nitrogen Management Consortium, and accepted through separate admini strative action by FDEP (acceptance of the 2002 RA, 2007 RA Update & 2009 RA Addendum) and EPA (establishment of the 1998 federally-recognized TMDL for Tampa Bay)

.

Bay Segment Tampa Bay NMC Proposed Alternative Total Nitrogen Load expressed as tons/year Tampa Bay NMC Proposed Total Phosphorus Load expressed as tons/year Old Tampa Bay 486 104 Hillsborough Bay 1,451 1,093 Middle Tampa Bay 799 140 Lower Tampa Bay 349 52 Remainder of Lower Tampa Bay 629 112


### 3.0 Tampa Bay TN and TP Criteria: Analyses and Results

Multiple analyses were completed in the evaluation of potential T N and TP criteria expressed as inbay concentrations. These include:

• examination of the relationships between TN and TP loadings to in-bay TN and TP concentrations, respectively;

• examination of relationships between monthly TN and TP concen trations with chlorophyll

a concentrations;

• examination of relationships between annual TN and TP concen trations with chlorophyll a concentrations; and

• application of a reference period approach to establishing T N and TP concentration-based criteria.
The data used in these analyses are defined in Attachment 2. Th e following describes these analyses and the results obtained.
3.1 Evaluation of Relationships Between In-bay TN and TP Concentr ations and TN and TP Loads
Since the current proposed TN and TP criteria are expressed as loads, the simplest method to propose criteria expressed as in-bay concentrations would be b ased on the potential relationships between in-bay TN and TP concentrations and TN and TP loa ds delivered to each segment. All four mainstem segments have annual TMDL TN loads recognized by bo th EPA and DEP. If significant relationships are found between the nutrient load s and their respective in-bay concentrations, then the proposed numeric nutrient criteri a loads could be expressed as concentrations.
Monthly segment-specific TN and TP concentrations were merged wi th monthly segment-specific TN and TP loads resulting in a dataset of monthly values of TN and TP concentrations and loads. Plots of these data were inspected, with TN and TP concentra tions as functions of TN and TP loads, respectively, including various lag and cumulative load effects. Graphical results for each segment16 February 2011 12

are provided in Attachment 3 (TN) and Attachment 4 (TP). The relationships examined are between the in-bay nutrient concentrations and the following:

• Current months load,

• Previous month load,

• Cumulative loads over current and previous month, and

• Cumulative loads over current and two previous months.
The graphical representations of the relationships were then u sed to guide evaluation of relationships between TN and TP concentrations and loads th at may explain large proportions of the monthly variation in concentrations.
No relationships were found between TN concentrations and pot ential TN load explanatory variables that explained more than 20% of the variation in T N concentrations in Hillsborough Bay or more than 10% of the variation in concentrations in th e other three bay segments (Table 4). In Old Tampa Bay, the natural log-transformed three-month cumulat ive TN load explained only 7% of the variation in in-bay TN concentrations. In Hillsborou gh Bay, the natural log-transformed twomonth cumulative TN load explained 33% of the variation in in-bay TN concentrations. In Middle Tampa Bay, 10% of the variation in in-bay TN concentration s was explained by the natural logtransformed three-month cumulative TN load, and 20% of the va riation in in-bay TN concentrations in Lower Tampa Bay was explained by the natural log-transformed cumulative threemonth TN load. This suggests that there are some in-bay processes (e.g., sedimentation, denitrification, and transport within the bay and exchan ge with the Gulf of Mexico) that affect the relationship between the TN loads and the resultant in-bay TN concentrations in Tampa Bay. The relationships between TP concentrations and potential TP loading explanatory variables that were found varied appreciably among bay segments (Table 3). In Old Tampa Bay, the natural logtransformed three-month cumulative TP load explained 44% of th e variation in TP concentrations. In Hillsborough Bay, the natural log-transformed current mont h TP load explained 33% of the variation in in-bay TP concentrations. In Middle Tampa Bay, 47% of the variation in TP concentrations was explained by the natural log-transformed three-month cumulative TP load, and 20% of the variation in TP concentrations in Lower Tampa Bay was explained by the natural logtransformed cumulative three-month TP load.
The results of these analyses do not provide adequate evidence to support recommendations for TN and TP concentration criteria based on the relationships b etween the in-bay nutrient concentrations and the nutrient loads to those segments.

Table 4 . Best

- fit regressions of TN and TP concentrat ions on TN and TP loads, respectively. Segment

– Variable Regression p > F r 2 Old Tampa Bay

– [TN]    = 0

. 19 + 0

. 112 ∗ ln Cumulative 3

- Month TN Load <0.0001 0.07 Old Tampa Bay

– [TP]    = − 0

. 32 + 0

. 15 ∗ ln Cumulative 3

- Month TP Load <0.0001 0.44 Hillsborou gh Bay

– [TN]    = − 0

. 25 + 0

. 189 ∗ ln Cumulative 2

- Month TN Load <0.0001 0.20 Hillsborough Bay

– [TP]    = 0

. 32 + 0

. 0008 ∗ ln Current Month TP Load <0.0001 0.33 Middle Tampa Bay

– [TN]    = 0

. 5 + 0

. 0005 ∗ ln Cumulative 3

- Month TN Load <0.0001 0.10 Middle Tamp a Bay

– [TP]    = − 0

. 08 + 0

. 089 ∗ ln Cumulative 3

- Month TP Load <0.0001 0.47 Lower Tampa Bay

– [TN]    = 0

. 3 + 0

. 036 ∗ ln Cumulative 3

- Month TN Load <0.0141 0.02 Lower Tampa Bay

– [TP]    = 0

. 1 + 0

. 0005 ∗ ln Cumulative 3

- Month TP Load <0.0001 0.2016 February 2011 13

3.2 Evaluation of Relationships Between Chlorophyll a and TN and TP Concentrations
The second data analysis approach examined the potential relat ionships between chlorophyll a concentrations and either TN or TP concentrations in each bay segment. Chlorophyll a thresholds have been established by the TBEP as part of the nitrogen management plan for the bay. If significant relationships are found, then the chlorophyll a thresholds could be used to determine the corresponding nutrient concentrations for use as numeric nutrient criteria. There are two temporal scales that can be examined – monthly and annual.
3.2.1 Evaluation of Relationships Between Monthly Chlorophyll a and TN and TP Concentrations
Initially, monthly segment-specific chlorophyll a and nutrient (TN and TP) concentrations were plotted. A series of variables based on the ambient TN and TP concentrations, including various lag concentrations, was examined. Graphical results are provi ded in Attachments 5 (TN) and 6 (TP) for each segment as scatter plots of chlorophyll a and the following:

• Current month TN and TP concentration,

• Previous month TN and TP concentration,

• Average of current and previous months TN and TP concentration s, and

• Average of current and two previous months TN and TP concentra tions,
The graphical representations of the relationships were then use d to guide evaluation of relationships between chlorophyll a and TN and TP concentrations that may explain the monthly variation in chlorophyll a concentrations.
No relationships were found between chlorophyll a and TN concentrations that explained more than 24% of the variation in chlorophyll a (maximum r 2 = 0.24) (Table 5). In Old Tampa Bay, the mean two-month TN concentration explained 12% of the variat ion in in-bay chlorophyll a concentrations. In Hillsborough Bay, the same-month TN concen tration explained 33% of the variation in in-bay chlorophyll a concentrations. In Middle Tampa Bay, 17% of the variatio n in inbay chlorophyll a concentrations was explained by the same-month TN concentra tion, and only 6% of the variation in in-bay chlorophyll a concentrations in Lower Tampa Bay was explained by the same-month TN concentration.
No relationships were found between chlorophyll a and TP concentrations that explained more than 27% of the variation in chlorophyll a concentrations (Table 5). In Old Tampa Bay, the samemonth TP concentration explained 15% of the variation in in-bay chlorophyll a concentrations. In Hillsborough Bay, the mean two-month TP concentration explain ed 17% of the variation in in-bay chlorophyll a concentrations. In Middle Tampa Bay and Lower Tampa, th e same-month TP concentration explained 27% and 12%,respectively, of the vari ation in in-bay chlorophyll a concentrations.16 February 2011 14

Table 5 . Best

- fit regressions of m onthly chlorophyll a concentrations on TN and TP concentrations. Segment – Variable Regression p > F r 2 Old Tampa Bay

– TN chl  = 3.78 + 7.34*Mean 2

- month TN Concentration <0.0001 0.12 Old Tampa Bay

– TP chl  = 5.51 + 14.94*Mean TP Concentration <0.0001 0.15 Hillsborough Bay

– TN chl  = 1.22 + 15.7* Mean TN Concentration <0.0001 0.24 Hillsborough Bay

– TP chl  = 6.82+15.65*Mean 2

- month TP Concentration <0.0001 0.17 Middle Tampa Bay

– TN chl  = 2

. 75 + 7.45* Mean TN Concentration <0.0001 0.17 Middle Tampa Bay

– TP chl  = 3

. 45 + 15.59*Mean TP Concentration <0.0001 0.27 Lower Tampa Bay

– TN chl  = 3.02 + 2.9* Mean TN Concentratio n <0.0003 0.06 Lower Tampa Bay

– TP chl  = 2

. 97 + 10.22*Mean TP Concentration <0.0001 0.12
The results of these analyses do not provide adequate evidence to su pport recommendations for TN and TP concentration criteria based on the relationships b etween the in-bay chlorophyll a concentrations and the in-bay TN and TP concentrations in these segments.
3.2.2 Evaluation of Relationships Between Annual Chlorophyll a and TN and TP Concentrations
The relationships between annual mean chlorophyll a and annual mean TN and TP concentrations were examined as a potential means of developing nutrient criteri a. The annual mean TN and TP concentrations were classified according to whether the ann ual chlorophyll a threshold in a given segment was met or not. Based on this classification, the TN and TP concentrations for those years in which the threshold was met could be compared to those TN a nd TP concentrations for those years when the threshold was exceeded (Figures 2-9).
The results of this analysis indicate that on an annual ba sis, there are differences in the in-bay TN concentrations during years in which the threshold chloroph yll a targets are met and those in which they are not met. This is particularly the case in Hillsborough Bay and Middle Tampa Bay. Similar differences were found between the TP concentrations in those years in which the threshold chlorophyll a targets are met and those in which they are not met. However, these differences are not as discrete as seen in the TN concentrations.16 February 2011 15

Figure 2. Comparison of TN concentrations in Old T ampa Bay in those years when the chlorophyll a thresholds were exceeded and those years when the c hlorophyll a thresholds were not exceeded.

Figure 3. Comparison of TN concentrations in Hills borough Bay in those years when the chlorophyll a thresholds were exceeded and those years when the c hlorophyll a thresholds were not exceeded.16 February 2011 16

Figure 4. Comparison of TN concentrations in Middl e Tampa Bay in those years when the chlorophyll a thresholds were exceeded and those years when the c hlorophyll a thresholds were not exceeded.

Figure 5. Comparison of TN concentrations in Lower Tampa Bay in those years when the chlorophyll a thresholds were exceeded and those years when the c hlorophyll a thresholds were not exceeded.16 February 2011 17

Figure 6. Comparison of TP concentrations in Old T ampa Bay in those years when the chlorophyll a thresholds were exceeded and those years when the c hlorophyll a thresholds were not exceeded.

Figure 7. Comparison of TP concentrations in Hills borough Bay in those years when the chlorophyll a thresholds were exceeded and those years when the c hlorophyll a thresholds were not exceeded.16 February 2011 18

Figure 8. Comparison of TP concentrations in Middl e Tampa Bay in those years when the chlorophyll a thresholds were exceeded and those years when the c hlorophyll a thresholds were not exceeded.

Figure 9. Comparison of TP concentrations in Lower Tampa Bay in those years when the chlorophyll a thresholds were exceeded and those years when the c hlorophyll a thresholds were not exceeded.16 February 2011 19

3.3 Reference Period TN and TP Concentrations as Concentration Crite ria
The analyses described above indicated that while there were signifi cant relationships between monthly nutrient concentrations and loads, and between month ly chlorophyll a and nutrient concentrations, these relationships explained very little of the observed variation in the dependent constituent. The evaluations of annual average chlorophyll a and annual TN and TP concentrations did provide evidence that for some segments, there were differences in nutrient concentrations between those periods in which chlorophyll a targets were met and those in which they were exceeded. Based on these findings, establishment of nutrient crit eria should be linked to the annual chlorophyll a and loading targets already established for Tampa Bay.
The fourth approach to developing concentration-based numer ic nutrient criteria is the reference period approach. Segment-specific chlorophyll a targets (values at this level or below indicate desirable conditions) have been previously established (Janicki a nd Wade, 1996). These targets were based on a 1992-1994 reference period. In 2000, a pr otocol for assessing whether the Tampa Bay segments were achieving these targets was developed (Janicki and Pri bble, 2000). This protocol, referred to as the Decision Matrix approach, con sidered the year-to-year variability in chlorophyll a concentrations and arrived at segment-specific chlorophyll a thresholds (values above this level indicate undesirable conditions). The threshold wa s the sum of the chlorophyll a target and 2X the standard error of the long-term chlorophyll a concentrations. FDEP has adopted these thresholds to assess compliance with the Tampa Bay Reasonable Assur ance.
Following this approach, numeric nutrient criteria for the four bay segments can be estimated. The 1992-1994 annual geometric mean concentrations are:

• Old Tampa Bay TN=0.75 mg/L TP=0.25 mg/L

• Hillsborough Bay TN=0.82 mg/L TP=0.37 mg/L

• Middle Tampa Bay TN=0.69 mg/L TP=0.24 mg/L

• Lower Tampa Bay TN=0.57 mg/L TP=0.08 mg/L
The following are the standard deviations of the mean annual TN and TP concentrations from the 1992-2009 period (which includes all years since the chloro phyll a concentration and TN loading targets were established):

• Old Tampa Bay TN=0.18 mg/L TP=0.06 mg/L

• Hillsborough Bay TN=0.19 mg/L TP=0.08 mg/L

• Middle Tampa Bay TN=0.18 mg/L TP=0.05 mg/L

• Lower Tampa Bay TN=0.17 mg/L TP=0.02 mg/L

In the same manner as the chlorophyll a thresholds were developed for Tampa Bay as described above, the proposed concentration-based TN and TP criteria are defined as the sum of the concentration targets and the standard deviation of the long -term mean annual TN and TP concentrations. Table 6 presents these proposed criteria.16 February 2011 20

Table 6. Proposed numeric nutrient criteria based on geometric mean of an nual concentrations for the period 1992-1994 and the standard deviat ion of the long-term TN and TP concentrations.

Segment TN Concentration (mg/L) TP Concentration (mg/L) Old Tampa Bay 0.9 3 0.3 1 Hillsborough Bay 1.0 1 0.45 Middle Tampa Bay 0.8 7 0.29 Lower Tampa Bay 0.7 4 0.10

The proposed TN and TP concentration criteria are compared to the observed geometric mean annual TN and TP concentrations in Figures 10 through 17. The horizontal lines represent the proposed criteria; the data to the right of the vertical li nes depict the TN and TP concentrations since the establishment of the reference period (1992-1994).

Figure 10. Comparison of proposed TN concentration criterion for Old Tampa Bay to the annual

geometric mean TN concentrations from 1981 through 2009.16 February 2011 21

Figure 11. Comparison of proposed TN concentration criterion for Hillsborough Bay to the annual

geometric mean TN concentrations from 1981 through 2009.

Figure 12. Comparison of proposed TN concentration criterion for Middle Tampa Bay to the annual

geometric mean TN concentrations from 1980 through 2009.16 February 2011 22

Figure 13. Comparison of proposed TN concentration criterion for Lower Tampa Bay to the annual

geometric mean TN concentrations from 1980 through 2009.

Figure 14. Comparison of proposed TP concentration criterion for Old Tampa Bay to the annual geometric mean TP concentrations from 1980 through 2009.16 February 2011 23

Figure 15. Comparison of proposed TP concentration criterion for Hillsborough Bay to the annual

geometric mean TP concentrations from 1980 through 2 009.

Figure 16. Comparison of proposed TP concentration criterion for Middle Tampa Bay to the annual

geometric mean TP concentrations from 1980 through 2 009.16 February 2011 24

Figure 17. Comparison of proposed TP concentration criterion for Lower Tampa Bay to the annual

geometric mean TP concentrations from 1980 through 2 009.



### 4.0 Conclusions

The following conclusions can be drawn from the analyses and results discussed above:

• Tampa Bay is nitrogen-limited as indicated by both ambient TN:T P ratios and nutrient addition bioassays.

• There is no discernable relationship between TN loadings and i n-bay TN concentrations or between TP loadings and in-bay TP concentrations in any bay se gment. This is not because loadings do not affect in-bay concentrations, but because va rious other confounding factors, for which sufficient data are not available, play a role i n relationships between loadings and concentrations

• On a monthly time scale, the relationships between either TN concen trations or TP concentrations and chlorophyll a concentrations do not explain a significant proportion o f the variability in the chlorophyll a concentrations to support development of concentrationbased numeric nutrient criteria in any bay segment.

• There are differences in TN concentrations and TP concentrat ions, particularly in Hillsborough Bay and Middle Tampa Bay, in those years when the chl orophyll a concentration thresholds are met when compared to those obser ved in years when the chlorophyll a concentration thresholds are not met. However, there is a great deal of variability within the data obtained in either group of yea rs and these differences are not16 February 2011 25

recommended as the basis for the establishment of concentration- based numeric nutrient criteria for Tampa Bay.

• The reference period approach is recommended for the establishmen t of concentrationbased TN and TP criteria for Tampa Bay. The segment-specific a nnual geometric mean nitrogen and phosphorus concentrations of the 1992-1994 pe riod were increased by one standard deviation (as derived from 1992-2009 data) to develo p the proposed concentration numeric nutrient criteria. These criteria are:
- Old Tampa Bay TN=0.93 mg/L TP=0.31 mg/L - Hillsborough Bay TN=1.01 mg/L TP=0.45 mg/L - Middle Tampa Bay TN=0.87 mg/L TP=0.29 mg/L - Lower Tampa Bay TN=0.74 mg/L TP=0.10 mg/L.



### 5.0 References

Borsuck, M.E., C.A.Stow, and K.E. Reckhow. 2004. Confoundin g effect of flow on estuarine response to nitrogen loading. Jour. Environ. Eng. 130:605-6 14.
Boynton, W.R. and W.M. Kemp. 2008. Estuaries. pp. 809-856. In: Nitrogen in the Marine Environment. 2nd Edition. Capone, D.G., Bronk, D.A., Mulho lland, M.R., and Carpenter, E.J. (eds.), Elsevier Inc., Burlington, Massachusetts.
Boynton, W.R., W.M. Kemp, and C.W. Keefe. 1982. A comparative analysis of nutrients and other factors influencing estuarine phytoplankton production. In : Estuarine Comparisons. Kennedy, V.S. (ed.). Academic Press, San Diego.
Chapra, S.C. 1997. Surface Water-Quality Modeling, McGra w-Hill, New York, N.Y.
Conley, D.J. 2000. Biogeochemical nutrient cycles and nutr ient management strategies. Hydrobiologia. 410:87-96.
Conley, D.J., H.W. Paerl, R.W. Howarth, D.F. Boesch, S.P. Seitzinger, K.E. Havens, C. Lancelot, and G.E. Likens. 2009. Controlling Eutrophication: Nitro gen and Phosphorus. Science. 323:1014- 1015.
Correll, D.L. 1999. Phosphorus: A rate limiting nutrient in surface waters. Poultry Science. 78:674-682.
Correll, D.L., and N.E. Tolbert. 1962. Ribonucleic acid -polyphosphate from algae. I. Isolation and physiology. Plant Physiol. 37:627-636.
D’Elia, C.F., J.G. Sanders, and W.R. Boynton. 1986. Nutri ent enrichment studies in a coastal plain estuary: Phytoplankton growth in large-scale continuous cul tures. Can. J. Fish. Aquat. Sci. 43:397- 406.16 February 2011 26

Elser, J.J., E.R. Marzolf, and C.R. Goldman. 1990. Phosph orus and nitrogen limitation of phytoplankton growth in the freshwaters of North America: a review and critique of experimental enrichments. Can. J. Fish. Aquat. Sci. 47:1468-1477.
Elser, J.J., M.E.S. Bracken, L.L Cleland, D.S. Gruner, W.S. harp ole, H. Hillebrand, J.T. Ngai, E.W. Seabloom, J.B. Shurin, and J.E. Smith. 2007. Global analysi s of nitrogen and phosphorus limitation of primary producers in freshwater, marine and terrestrial ecos ystems. Ecology Letters 10, doi: 10.1111/j.1461-0248.2007.01113x.
FDEP. 2002. Unpublished guidelines for determining nutrient li mitations in waterbodies under the Impaired Waters Rule. Watershed Assessment Division. Tallahassee.
Fisher, T.R., E.R. Peele, J.W. ammerman, and L.W. Harding, Jr. 19 92. Nutrient limitation of phytoplankton in Chesapeake Bay. Mar. Ecol. Prog. Ser. 82:51-63

. Hagy, J.D., L.P. Sanford, and W.R. Boynton. 2000. Estimatio n of net physical transport and hydraulic residence times for a coastal plain estuary using box models. Estuaries. 23:328-340.
Havens, K.E., and J. DeCosta. 1986. A comparison of phytopl ankton responses to nutrient additions in acidic and circumneutral pH lakewater. Hydr obiologia. 137:211-222.
Hecky, R.E., and P. Kilham. 1988. Nutrient limitation of p hytoplankton in freshwater and marine environment: A review of recent evidence on the effects of enrichment . Limnol. Oceanogr. 33:796-822.
Howarth, R.W. 1988. Nutrient limitation of net primary p roductivity in marine ecosystems. Annual Review of Ecology and Systematics. 19:89-110.

Howarth, R. W. 2008. Coastal nitrogen pollution: A revi ew of sources and trends globally and regionally. Harmful Algae. 8:14-20.
Howarth, R.W., and R. Marino. 2006. Nitrogen as the li miting nutrient for eutrophication in coastal marine ecosystems: Evolving views over three decades. Limnol. Oceanogr . 51:364-376.
Howarth, R.W., R. Marino, J. Land, and J.J. Cole. 198 8a. Nitrogen fixation in freshwater, estuarine, and marine ecosystems. 1. Rates and importance. Limnol. Ocea nogr. 33:669-687.
Howarth, R.W., R. Marino, and J.J. Cole. 1988b. Nit rogen fixation in freshwater, estuarine, and marine ecosystems. 2. Biogeochemical controls. Limnol. Oceanogr. 33:68 8-701.
Jahnke, J., H.J. Rick, and L. Aletsee. 1986. On the light an d temperature dependence of the minimum and maximum phosphorus contents in cells of the marine pla nkton diatom Thalassiosira rotula Meunier. J. Plankton Res. 8:549-555.
Janicki, A.J., and J.R. Pribble. 2000. Establishing a pr ocess for tracking chlorophyll-a concentrations and light attenuation in Tampa Bay. Tampa Ba y National Estuary Program Technical Publication #04-00.16 February 2011 27

Janicki, A., and D. Wade. 1996. Estimating critical extern al nitrogen loads for the Tampa Bay Estuary: An empirically based approach to setting management target s. Tampa Bay National Estuary Program Technical Publication #06-96.

Johansson, J.O.R. 2009. Nutrient enrichment studies of nat ural phytoplankton populations in Tampa Bay: A summary of results June 1993 to August 2009. City of Tampa Wastewater Department Bay Study Group.

Lee, Y.S., T. Seiki, T. Mukai, K. Takimoto, and M. Okada. 1996. Limiting nutrients of phytoplankton community in Hiroshima Bay, Japan. Water Res. 30: 1490-1494.
Lynch, M., and J. Shapiro. 1981. Predation, enrichment, a nd phytoplankton community structure. Limnol. Oceanogr. 26:86-102.
Malone, T.C., D.J. Conley, T.R. Fisher, P.M. Glibert, L.W. Harding, and K.G. Sellner. 1996. Scales of Nutrient-Limited Phytoplankton Productivity in Chesapeake Bay. Estuaries. Vol. 19, No. 2. Dedicated Issue: Nutrients in Coastal Waters. pp. 371-385.

McComb, A.J., R.P. Atkins, P.B. Birch, and R.J. Lukatelich. 1981. Eutrophication in the PeelHarvey estuarine system, Western Australia. In: Estuaries and Nutr ients. Nielsen, B.J., and L.E. Cronin (eds). Humana Press. Clifton, NJ, p 569-582.

Monsen, N.E., J.E. Cloern, and L.V. Lucus. 2002. A comment o n the use of flushing time, residence time, and age as transport time scales. Limnol. Oceanogr. 47:154 5-1553.
National Research Council (NRC), 2000. Clean Coastal Waters: Understanding and Reducing the Effects of Nutrient Pollution. National Academy Press. Washingt on, D.C.
National Research Council (NRC), 1993. Managing wastewater i n coastal urban areas. National Academy Press. Washington, D.C.

Nixon, S.W., et al. (15 co-authors). 1996. The fate of nitrogen and phosphorus at the land-sea margin of the North Atlantic Ocean. Biogeochemistry. 35:141- 180.
Oviatt, C.A., A.A. Keller, P.A. Sampou, and L.L. Beatty. 1986. Patterns of productivity during eutrophication: a mesocosm experiment. Mar. Ecol. Prog. Ser. 28 :69-80.
Parsons, T.R., Y. Maita, and C.M. Lalli. 1984. A manua l of chemical and biological methods for seawater analysis. Pergamon Press.
Pennock, J.R., and J.H. Sharp. 1994. Temporal alternatio n between light- and nutrient-limitation of phytoplankton production in a coastal plain estuary. Mar . Ecol. Prog. Ser. 111:275-288.

Perez, E.A.A., J. DeCosta, and K.E. Havens. 1994. The effects of nutrient addition and pH manipulation in bag experiments on the phytoplankton of a smal l acidic lake in West Virginia, USA. Hydrobiologia. 291:93-103.16 February 2011 28

Redfield, A.C. 1934. On the proportion of organic derivat ives in sea water and their relation to the composition of plankton. James Johnstone Memorial Volume. Liver pool University Press. pp. 176-192.
Redfield, A.C. 1958. The biological control of chemical factors in the environment. Am. Sci. 46:205-222.
Ryther, J.H., and W.M. Dunstan. 1971. Nitrogen, phospho rus, and eutrophication in the coastal marine environment. Science. 171:1008–1013.
Schindler, D.W. 1974. Eutrophication and recovery in exper imental lakes: Implications for lake management. Science. 184:897-899.
Schindler, D.W. 1975. Whole-lake eutrophication experiment s with phosphorus, nitrogen and carbon. Verh. Int. Verein. Limnol. 19:3221-3231.
Schindler, D.W. 1977. The evolution of phosphorus limitat ions in lakes. Science. 195:260-262.
Shapiro, J. 1980. The importance of trophic-level interact ions to the abundance and species composition of algae in lakes. In: Hypertrophic Ecosystems, Developm ents in Hydrobiology. Barica, J. and L.R. Mur (eds). Junk, The Netherlands Vol. 2 . pp.105-116.
Sicko-Goad, L., and T.E. Jensen. 1976. Phosphate metabolism in blue-green algae. II. Changes in phosphate distribution during starvation and the “polyphosp hate overplus” phenomenon in Plectonema boryanum . Am. J. Bot. 63:183-188.
Smith, S.V. 1984. Phosphorus versus nitrogen limitation in the marine environment. Limnol. Oceanogr. 29:1149-1160.

Tampa Bay Nitrogen Management Consortium (TBNMC). 2010. Letter to Water Docket, U.S. Environmental Protection Agency, March 8, 2010.
Taylor, D., S. Nixon, S. Granger, and B. Buckley. 1995. Nu trient limitation and the eutrophication of coastal lagoons. Mar. Ecol. Prog. Ser. 127:235-244.
Terry, K.L., J. Hirate, and E.A. Laws. 1985. Light-, nitrogen -, and phosphorus-limited growth of Phaeodactylum tricornutum Bohlin strain TFX-1: chemical composition, carbon partitio ning, and the diel periodicity of physiological processes. J. Exp. Mar. Biol. Ecol. 86:85-100.
Thomas, W. H. 1970a. On nitrogen deficiency in tropical p arameters in poor and rich water. Limnol. Oceanogr. 15:380-385.

Thomas, W. H. 1970b. Effect of ammonium and nitrate conc entration on chlorophyll increases in natural tropical Pacific phytoplankton populations. Li mnol. Oceanogr. 15:386-394.
Wynne, D., and G.Y. Rhee. 1986. Effects of light intensity and quality on the relative N and P requirement (the optimum N:P ratio) of marine planktonic alga e. J. Plankton Res. 8:91-103.Attachment 1 Results of City of Tampa Bay Study Group Bioassays (Johansson, 2009)

## NUTRIENT ENRICHMENT STUDIES OF NATURAL PHYTOPLANKTON POPULATIONS IN TAMPA BAY

A SUMMARY OF RESULTS JUNE 1993 TO AUGUST 2009

J.O.R. JOHANSSON THE CITY OF TAMPA WASTEWATER DEPARTMENT BAY STUDY GROUP

NOVEMBER 16, 2009

### NUTRIENT ENRICHMENT STUDIES OF NATURAL PHYTOPLANKTON POPULATIONS IN TAMPA BAY. A SUMMARY OF RESULTS

JUNE 1993 TO 

## AUGUST 2009

INTRODUCTION

The Bay Study Group (BSG) has been conducting nutrient enrichment studies (bioassays) on natural phytoplankton populations at four locations in Tampa Bay since 1993 (Figure 1). This report will summarize findings for all tests that have been performed to date, from June 1993 through August 2009.

Figure 1. Phytoplankton nutrient bioassay monitoring stations in Tampa Bay.

METHODS

The bioassays have been performed on the natural phytoplankton population collected from surface waters of the four stations, with one station located in each of the four major segment of Tampa Bay, as shown below:

 Hillsborough Bay - COT4

 Old Tampa Bay - COT40

 Middle Tampa Bay - COT13

 Lower Tampa Bay - COT95

### Bioassay measurements were performed on a quarterly schedule from June 1993 through August

1995 for all stations except COT95. At this station monthly tests were conducted during the first year of the program. From February 1996 to the present, bioassays have been performed twi ce per year, during late winter and late summer.

The bioassay method used is similar to a method used in Chesapeake Bay waters (see Fisher et al. 1992a and b). An outline of the specific method used by the BSG is provided here.

A large volume of surface water is collected for the following nutrient treatments. Each treatment is conducted in duplicate on 3l samples:

 Controls (no nutrient addition).

 N-additions (NH3-N added to reach a final concentration in the sample of near 50uM).

 P-additions (PO4- P added to reach a final concentration in the sample of near 5uM).

 N+P-additions (combination of the N-additions and P-additions).

The treatment samples are incubated under natural sunlight in ambient bay water temperatures for an appropriate incubated period (usually 48h for the late winter and 24h for the late summer tests).

The growth response of the natural phytoplankton community to the different treatments is determined through measurements in changes of algal biomass, measured as chlorophyll-a.

Pa ired t-test statistics (p<0.05) and non-statistical evaluations of treatment responses are used to interpret the bioassay results and to group the growth response to the nutrient additions into the following response categories:

 Exclusive N limitation: (1) the addition of P induced no response relative the control, and (2) the addition of N alone had virtually the same effect as the addition of N+P.

 Primary N limitation: (1) the addition of P alone induced little response relative the control, (2) the addition of N alone induced a response, and (3) the addition of N+P induced the largest response.

 Balanced NP limitation: (1) the addition of N and P alone induced no response relative the control, (2) the addition of N+P induced a large response.

 Exclusive P limitation: (1) the addition of N induced no response relative the control, and (2) the addition of P alone had virtually the same effect as the addition of N+P.

 Primary P limitation: (1) the addition of N alone induced little response relative the control, (2) the addition of P alone induced a response, and (3) the addition of N+P induced the largest response.

 No response to any nutrient addition, indicating nutrient saturation, light limitation, and/or insufficient incubation time.

### RESULTS

A total of 152 bioassay experiments have been conducted since the start of the program in June 1993.

The growth response of the natural phytoplankton community to the different nutrient treatments were grouped into the response categories described above. The result of these analyses is summarized in Table 1.

Table 1. Results from natural phytoplankton nutrient bioassays in the four major subsections of Tampa Bay, 1993

– 2009. Bioassay response Hillsborough Bay COT4 Old Tampa Bay

COT40 Middle Tampa Bay

COT13 Lower Tampa Bay

COT95 Exclusive N limitation 29 25 28 39 Primary N limitation 2 2 2 1 Balanced 0 0 0 0 Exclusive P limitation 0 0 0 0 Primary P limitation 0 0 0 0 No response 0 3 0 0



### DISCUSSION AND CONCLUSION

One-hundred-fifty-two natural phytoplankton community nutrient limitation experiments have been conducted in Tampa Bay by the BSG from 1993 to the present. The strong nitrogen dependence by the Tampa Bay phytoplankton community is obvious in all four bay segments; of the 152 bioassay tests conducted to date, 149 indicate that nitrogen was the stronger limiting nutrient. None of the 152 tests have indicated that phosphorous was the stronger limiting nutrient. However, three tests in OTB have shown a lack of phytoplankton growth response by either N or P additions.

REFERENCES

Fisher, T.R., A.B. Gustafson, K.G. Sellner, and R.B. Lacouture.1992a. Nutrient bioassays in Chesapeake Bay to assess nutrients limiting algal growth. Progress Rep. MD Dept. of the Environment, Jan. 1992, Baltimore, MD.

Fisher, T.R., E.R. Peele, J.W. Ammerman, and L.W. Harding, Jr. 1992b. Nutrient limitation of phytoplankton in Chesapeake Bay. Mar. Ecol. Prog. Ser., 82:51-63.Attachment 2 Data SourcesThe following data sources were used in the analyses performed for this task:

 The water quality data were obtained from the Environmental Protection Commission of Hillsborough County (EPCHC). Their samplin g program collects surface samples on a monthly basis from the following series of fixed stations. Among others, the primary analytes included:

- Chlorophyll a

- TN

- TP

- Salinity.

Old Tampa Bay 46 60 64 47 65 63 66 40 41 67 50 38 51 68 36 Hillsborough Bay 44 70 52 6 71 7 55 08 73 80 Middle Tampa Bay 11 9 81 84 33 32 13 14 82 28 16 19 Lower Tampa Bay 25 23 21 91 90 24 95 92 93

 The TN and TP loading estimates were obtained from a series of reports produced for the TBEP (1985-2002) and FDEP (2003-2007). These are:

Zarbock, H., A. Janicki, D. Wade, D. He imbuch, and H. Wilson. 1994. Estimates of Total Nitrogen, Total Phosphorus, and Total Suspended Solids Loadings to Tampa Bay, Florida. Technical Publication #0 4-94 of the Tampa Bay National Estuary Program. Prepared by Coastal Envi ronmental, Inc. Prepared for Tampa Bay National Estuary Program. St. Petersburg, FL.

Zarbock, H.W., A.J. Janicki, and S.S. Janicki. 1996. Estimates of Total Nitrogen, Total Phosphorus, and Total Suspended Solids Loadings to Tampa, Bay, Florida. Technical Appendix: 1992-94 Tot al Nitrogen Loadings to Tampa Bay. Technical Publication #19-96 of the Tampa Bay National Estuary Program. Prepared by Coastal Environmental, Inc. Prepared for Tampa Bay National Estuary Program. St. Petersburg, FL.Pribble, R., A. Janicki, H. Zarbock, S. Janicki, and M. Winowitch. 2001. Estimates of Total Nitrogen, Total Phosphorus, Total Suspended Solids, and Biochemical Oxygen Demand Loadings to Tampa, Bay, Florid a: 1995-1998. Technical report #05-01 of the Tampa Bay Estuary Program. Prepared by Janicki Environmental, Inc. Prepared for Tampa Bay Estuary Program. St. Petersburg, FL. Poe, A., K. Hackett, S. Janicki, R. Pribble, and A. Janicki. 2005. Estimates of Total Nitrogen, Total Phosphorus, Total Suspended Solids, and Biochemical Oxygen Demand Loadings to Tampa Bay, Florid a: 1999-2003. Technical Report #02-05 of the Tampa bay Estuary Program. Prepared by Janicki Environmental, Inc. Prepared for Tampa Bay National Estuary Program. St. Petersburg, FL.

Janicki Environmental, Inc. 2008. Estimat es of Total Nitrogen, Total Phosphorus, Total Suspended Solids, and Biochemical Oxygen Demand Loadings to Tampa Bay, Florida: 2004-2007. Prepared by Janicki Environmental, Inc. Prepared for Florida Department of Environmental Protections. Tallahassee, FL.Attachment 3 Monthly TN Concentrations and Monthly TN Loads

## Old Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

Current Month TN Load (tons/month) 0 100 200 300



## Hillsborough Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

Current Month TN Load (tons/month) 0 100 200 300 400 500 600 700 800 900 1000

## Middle Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6

Current Month TN Load (tons/month) 0 100 200 300 400 500 600



## Lower Tampa Bay Monthly Mean TN Concentration (mg/L)

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

Current Month TN Load (tons/month) 0 100 200 300 400 500 600 700 800 900

## Old Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

Previous Month TN Load (tons/month) 0 100 200 300



## Hillsborough Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

Previous Month TN Load (tons/month) 0 100 200 300 400 500 600 700 800 900 1000

## Middle Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6

Previous Month TN Load (tons/month) 0 100 200 300 400 500 600



## Lower Tampa Bay Monthly Mean TN Concentration (mg/L)

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

Previous Month TN Load (tons/month) 0 100 200 300 400 500 600 700 800 900

## Old Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

Cumulative Two-Month TN Load (tons) 0 100 200 300 400 500



## Hillsborough Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

Cumulative Two-Month TN Load (tons) 0 200 400 600 800 1000 1200 1400 1600 1800 2000

## Middle Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6

Cumulative Two-Month TN Load (tons) 0 100 200 300 400 500 600 700 800 900



## Lower Tampa Bay Monthly Mean TN Concentration (mg/L)

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

Cumulative Two-Month TN Load (tons) 0 100 200 300 400 500 600 700 800 900 1000 1100

## Old Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

Cumulative Three-Month TN Load (tons) 0 100 200 300 400 500 600



## Hillsborough Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

Cumulative Three-Month TN Load (tons) 0 1000 2000 3000

## Middle Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6

Cumulative Three-Month TN Load (tons) 0 100 200 300 400 500 600 700 800 900



## Lower Tampa Bay Monthly Mean TN Concentration (mg/L)

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

Cumulative Three-Month TN Load (tons) 0 200 400 600 800 1000 1200 1400

## Old Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

ln Current Month TN Load (tons/month) 1234 5 6



## Hillsborough Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

ln Current Month TN Load (tons/month) 3 4 5 6 7

## Middle Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6

ln Current Month TN Load (tons/month) 1234 5 6 7



## Lower Tampa Bay Monthly Mean TN Concentration (mg/L)

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

ln Current Month TN Load (tons/month) 1234 5 6 7

## Old Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

ln Cumulative Two-Month TN Load (tons) 234 5 6 7



## Hillsborough Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

ln Cumulative Two-Month TN Load (tons) 4 5 6 7 8

## Middle Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6

ln Cumulative Two-Month TN Load (tons) 234 5 6 7



## Lower Tampa Bay Monthly Mean TN Concentration (mg/L)

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

ln Cumulative Two-Month TN Load (tons) 234 5 6 7

## Old Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

ln Cumulative Three-Month TN Load (tons) 3 4 5 6 7



## Hillsborough Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

ln Cumulative Three-Month TN Load (tons) 4 5 6 7 8

## Middle Tampa Bay Monthly Mean TN Concentration (mg/L)

0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6

ln Cumulative Three-Month TN Load (tons) 3 4 5 6 7



## Lower Tampa Bay Monthly Mean TN Concentration (mg/L)

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

ln Cumulative Three-Month TN Load (tons) 3 4 5 6 7 8Attachment 4 Monthly TP Concentrations and Monthly TP Loads

## Old Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

Current Month TP Load (tons/month) 0 10 20 30 40 50 60 70 80 90 100 110 120 130



## Hillsborough Bay Monthly Mean TP Concentration (mg/L)

0.00 1.00 2.00 3.00

Current Month TP Load (tons/month) 0 1000 2000 3000

## Middle Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80

Current Month TP Load (tons/month) 0 100 200 300



## Lower Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60

Current Month TP Load (tons/month) 0 100 200 300

## Old Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

Previous Month TP Load (tons/month) 0 10 20 30 40 50 60 70 80 90 100 110 120 130



## Hillsborough Bay Monthly Mean TP Concentration (mg/L)

0.00 1.00 2.00 3.00

Previous Month TP Load (tons/month) 0 1000 2000 3000

## Middle Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80

Previous Month TP Load (tons/month) 0 100 200 300



## Lower Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60

Previous Month TP Load (tons/month) 0 100 200 300

## Old Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

Cumulative Two-Month TP Load (tons) 0 20 40 60 80 100 120 140 160 180 200



## Hillsborough Bay Monthly Mean TP Concentration (mg/L)

0.00 1.00 2.00 3.00

Cumulative Two-Month TP Load (tons) 0 1000 2000 3000 4000

## Middle Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80

Cumulative Two-Month TP Load (tons) 0 100 200 300 400



## Lower Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60

Cumulative Two-Month TP Load (tons) 0 100 200 300 400 500

## Old Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

Cumulative Three-Month TP Load (tons) 0 20 40 60 80 100 120 140 160 180 200 220



## Hillsborough Bay Monthly Mean TP Concentration (mg/L)

0.00 1.00 2.00 3.00

Cumulative Three-Month TP Load (tons) 0 1000 2000 3000 4000

## Middle Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80

Cumulative Three-Month TP Load (tons) 0 100 200 300 400



## Lower Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60

Cumulative Three-Month TP Load (tons) 0 100 200 300 400 500

## Old Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

ln Current Month TP Load (tons/month) 1234 5



## Hillsborough Bay Monthly Mean TP Concentration (mg/L)

0.00 1.00 2.00 3.00

ln Current Month TP Load (tons/month) 3 4 5 6 7 8

## Middle Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80

ln Current Month TP Load (tons/month) -1 0 1 2 3 4 5 6



## Lower Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60

ln Current Month TP Load (tons/month) 01234 5 6

## Old Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

ln Cumulative Two-Month TP Load (tons) 234 5 6



## Hillsborough Bay Monthly Mean TP Concentration (mg/L)

0.00 1.00 2.00 3.00

ln Cumulative Two-Month TP Load (tons) 4 5 6 7 8 9

## Middle Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80

ln Cumulative Two-Month TP Load (tons) 01234 5 6



## Lower Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60

ln Cumulative Two-Month TP Load (tons) 1234 5 6

## Old Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

ln Cumulative Three-Month TP Load (tons) 234 5 6



## Hillsborough Bay Monthly Mean TP Concentration (mg/L)

0.00 1.00 2.00 3.00

ln Cumulative Three-Month TP Load (tons) 4 5 6 7 8 9

## Middle Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80

ln Cumulative Three-Month TP Load (tons) 1234 5 6



## Lower Tampa Bay Monthly Mean TP Concentration (mg/L)

0.00 0.10 0.20 0.30 0.40 0.50 0.60

ln Cumulative Three-Month TP Load (tons) 1234 5 6 7Attachment 5 Monthly Chlorophyll a Concentrations and Monthly TN Concentrations

## Old Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Monthly Mean TN (mg/L) 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1



## Hillsborough Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0 60.0

Monthly Mean TN (mg/L) 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

## Middle Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Monthly Mean TN (mg/L) 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6



## Lower Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0

Monthly Mean TN (mg/L) 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

## Old Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Previous Month TN Concentration (mg/L) 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1



## Hillsborough Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0 60.0

Previous Month TN Concentration (mg/L) 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1

## Middle Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Previous Month TN Concentration (mg/L) 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6



## Lower Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0

Previous Month TN Concentration (mg/L) 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3

## Old Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Average Two-Month TN Concentration (mg/L) 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7



## Hillsborough Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0 60.0

Average Two-Month TN Concentration (mg/L) 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9

## Middle Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Average Two-Month TN Concentration (mg/L) 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5



## Lower Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0

Average Two-Month TN Concentration (mg/L) 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0

## Old Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Average Three-Month TN Concentration (mg/L) 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6



## Hillsborough Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0 60.0

Average Three-Month TN Concentration (mg/L) 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7

## Middle Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Average Three-Month TN Concentration (mg/L) 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3



## Lower Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0

Average Three-Month TN Concentration (mg/L) 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0Attachment 6 Monthly Chlorophyll a Concentrations and Monthly TP Concentrations

## Old Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0

Monthly Mean TP (mg/L) 0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70



## Hillsborough Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0 60.0

Monthly Mean TP (mg/L) 0.00 1.00 2.00 3.00

## Middle Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Monthly Mean TP (mg/L) 0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80



## Lower Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0

Monthly Mean TP (mg/L) 0.00 0.10 0.20 0.30 0.40 0.50 0.60

## Old Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0

Previous Month TP Concentration (mg/L) 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7



## Hillsborough Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0 60.0

Previous Month TP Concentration (mg/L) 0123

## Middle Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Previous Month TP Concentration (mg/L) 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8



## Lower Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0

Previous Month TP Concentration (mg/L) 0.0 0.1 0.2 0.3 0.4 0.5 0.6

## Old Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0

Average Two-Month TP Concentration (mg/L) 0.0 0.1 0.2 0.3 0.4 0.5 0.6



## Hillsborough Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0 60.0

Average Two-Month TP Concentration (mg/L) 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6 1.7

## Middle Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Average Two-Month TP Concentration (mg/L) 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7



## Lower Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0

Average Two-Month TP Concentration (mg/L) 0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.35 0.40 0.45

## Old Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0

Average Three-Month TP Concentration (mg/L) 0.05 0.10 0.15 0.20 0.25 0.30 0.35 0.40 0.45 0.50 0.55 0.60



## Hillsborough Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0 40.0 50.0 60.0

Average Three-Month TP Concentration (mg/L) 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4

## Middle Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 10.0 20.0 30.0

Average Three-Month TP Concentration (mg/L) 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7



## Lower Tampa Bay Monthly Mean Chlorophyll a (ug/L)

0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0

Average Three-Month TP Concentration (mg/L) 0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.35 0.40